Rapid prototyping for high-pressure microfluidics

Rein, Carlie; Toner, Mehmet; Sevenler, Derin

doi:10.1038/s41598-023-28495-2

Cited by 13 publications

(6 citation statements)

References 12 publications

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“…This unique respiratory property makes them ideal for long-term studies in sealed chambers. Our results confirm the practicality of growing these cells in large, sealed chambers (minimum diameter: 10 mm), which simplifies experimental setups and increases the practicality of the microfluidic platform (38). Additionally, we observed a morphological change in the shape of SH-SY5Y cells during cell division, from stretched to spherical.…”

Section: Discussionsupporting

confidence: 79%

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Proliferation of SH-SY5Y neuroblastoma cells on confined spaces

Kalwarczyk,

Lukasiak,

Woznica

et al. 2024

Preprint

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This study investigated the proliferation dynamics of SH-SY5Y neuroblastoma cells in a confined environment. The effects of seeding concentrations, culture surfaces and long-term maintenance in a sealed chamber were systematically investigated. Seeding concentration experiments revealed a clear correlation between initial cell number and proliferation rate, highlighting the importance of precise adjustments for optimal cell culture results. The study of available surface effects showed a nuanced relationship between chamber dimensions and cell growth, with confluence stability observed in larger chambers with diameters greater than 10 mm. The study also compared SH-SY5Y cells with HeLaGFP and 16HBE14𝜎 cells, revealing growth patterns influenced by chamber size. A properly designed sealed chamber with continuous real-time microscopic monitoring showed comparable cell growth to standard culture chambers. The research provides valuable insights into optimising SH-SY5Y cell culture practices, particularly in microfluidic systems.

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Section: Discussionsupporting

confidence: 79%

“…Observations support the practicality of culturing these cells in large, sealed plastic chambers (minimum diameter: 10 mm). The preference for rigid polymers over softer polymers, such as PDMS with higher gas permeability, streamlines experimental setups and increases the practicality of the microfluidic platform(33).…”

Section: Discussionmentioning

confidence: 99%

Proliferation of SH-SY5Y neuroblastoma cells on confined spaces

Kalwarczyk,

Lukasiak,

Woznica

et al. 2024

Preprint

View full text Add to dashboard Cite

show abstract

“…A channel geometry with a large cross sectional aspect ratio and high curvature was designed to maximize focusing throughput, by both increasing the channel cross section and also delaying the breakdown of primary Dean vortices at higher flow rates 43 . These devices were fabricated from rigid epoxy bonded to glass to prevent channel inflation under high operating pressures (described in “Methods”) 44 . We assessed the focusing performance for Jurkat cells suspended in PBS with HA across a range of flow rates, HA concentrations, and channel heights.…”

Section: Resultsmentioning

confidence: 99%

High throughput intracellular delivery by viscoelastic mechanoporation

Sevenler,

Toner

2024

Nat Commun

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Brief pulses of electric field (electroporation) and/or tensile stress (mechanoporation) have been used to reversibly permeabilize the plasma membrane of mammalian cells and deliver materials to the cytosol. However, electroporation can be harmful to cells, while efficient mechanoporation strategies have not been scalable due to the use of narrow constrictions or needles which are susceptible to clogging. Here we report a high throughput approach to mechanoporation in which the plasma membrane is stretched and reversibly permeabilized by viscoelastic fluid forces within a microfluidic chip without surface contact. Biomolecules are delivered directly to the cytosol within seconds at a throughput exceeding 250 million cells per minute. Viscoelastic mechanoporation is compatible with a variety of biomolecules including proteins, RNA, and CRISPR-Cas9 ribonucleoprotein complexes, as well as a range of cell types including HEK293T cells and primary T cells. Altogether, viscoelastic mechanoporation appears feasible for contact-free permeabilization and delivery of biomolecules to mammalian cells ex vivo.

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“…Master molds were fabricated by photolithography of SU-8 silicon wafers, and patterns transferred to a poly(dimethyl) siloxane replica mold used for epoxy casting, as described in detail elsewhere. 50 1.5 cm segments of 1/16 th inch ID polyethylene tubing were integrated for the device inlets and outlet during epoxy casting.…”

Section: Methodsmentioning

confidence: 99%

High throughput intracellular delivery by viscoelastic mechanoporation

Sevenler

Toner

2023

Preprint

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Brief and intense electric fields (electroporation) and/or tensile stresses (mechanoporation) have been used to temporarily permeabilize the plasma membrane of mammalian cells for the purpose of delivering materials to the cytosol. However, electroporation can be harmful to cells, while efficient mechanoporation strategies have not been scalable due to the use of narrow constrictions or needles which are susceptible to clogging. Here we report a method of mechanoporation in which cells were stretched and permeabilized by viscoelastic flow forces without surface contact. Inertio-elastic cell focusing aligned cells to the center of the device, avoiding direct contact with walls and enabling efficient (95%) intracellular delivery to over 200 million cells per minute. Functional biomolecules such as proteins, RNA, and ribonucleoprotein complexes were successfully delivered to Jurkat cells. Efficient intracellular delivery to HEK293T cells and primary activated T cells was also demonstrated. Contact-free mechanoporation using viscoelastic fluid forces appears to be feasible method for efficient and high throughput intracellular delivery of biomolecules to mammalian cells ex vivo.

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Rapid prototyping for high-pressure microfluidics

Cited by 13 publications

References 12 publications

Proliferation of SH-SY5Y neuroblastoma cells on confined spaces

Proliferation of SH-SY5Y neuroblastoma cells on confined spaces

High throughput intracellular delivery by viscoelastic mechanoporation

High throughput intracellular delivery by viscoelastic mechanoporation

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